Tissue Antigens (1977), 10, 291-302 Published by Munksgaard, Copenhagen, Denmark No part may be reproduced by any process without written permission from the authods)
Histocompatibility Determinants in Israeli Jewish Patients with Multiple Sclerosis Chaim Brautbar' , isac Cohen' , Esther Kahana', Milton Alter3, Fritz Jbrgensen4 and Lars Lamm4 Department of Immunology and Laboratory of Immunohematology, Hadassah Hebrew University Medical Center, Jerusalem, Israel; Department of Neurology, Hadassah Hebrew University Medical Center, Jerusalem, Israel; Department of Neurology, Temple University Health Sciences Center, Philadelphia, U.S.A. and Blood Grouping and Tissue Typing Laboratory, The University Hospital, Arhus, Denmark
'
The distribution of 24 HLA antigens of the A and B loci was investigated in 197 Israeli Jewish patients with multiple sclerosis (MS) from various Jewish ethnic origins including central and eastern Europe, countries bordering the Mediterranean, the Middle East and from native-born Israelis. The results were compared with the HLA antigen frequencies in a control sample of 455 unrelated individuals representing the general Jewish population. The frequency of HLA-Bw40 among all MS patients (15%) was significantly greater (P < 0.001) than among the controls (7%). In contrast to the findings in MS patients from other populations, there was no increased frequency of A3 and 07 and Dw2 was present in only one out of 28 patients. The study showed a similar distribution of HLA-A and -B locus antigens, especially of Bw40, in Jews of diverse ethnic origins represented in the control group.
Received for publication 1 1 March, revised, accepted 26 April 1977
Although an environmental factor has been suggested as the most likely cause of multiple sclerosis (MS) (Acheson 1972, Leibowitz & Alter 1973) the disease occurs 12 to 20 times more frequently within the family of a patient than in the general population (Berry 1969). Therefore, a genetic factor may also play a role in etiology. Numerous studies have shown as association between MS and certain histocom-
patibility (HLA) determinants, notably HLA-A3 of the first segregant series and B7 of the second series, (Bertram & Kuwert 1972, Naito et al. 1972, Jersaid et al. 1973a, Arnason et al. 1974). In a recent study it was clearly shown that HLA-B7 is the antigen with the greatest overall frequency (Jersild et al. 1975). Since it became possible to define and characterize lymphocytedefined determinants utilizing HLA-D homozygous typing
This work was supported, in part, by the Kroc Foundation for Medical Research, the Danish Medical Research Council, the Israel Multiple Sclerosis Society and Minktry of Health, and b y a special research grant kindly donated by Mr. and Mrs. Forrest Shaklee.
29 2
BRAUTBAR ET AL.
cells (Dupont et al. 1973, Jbrgensen et al. 1973) an HLA-B7 associated D locus determinant has been sought which was even more closely associated with MS. Indeed, Jersild et al. (1973b) were able to show a higher frequency of the lymphocyte-defined determinant Dw2 (formerly LD-7a) in MS patients (70%) than in the general population (16%). The presence of Dw2 was also apparently associated with a more malignant course of the disease (Jersild et al. 1973b). Further studies have confirmed that the frequency of Dw2 among MS patients is significantly higher than HLA-B7 (Jersild et al. 1975). In fact, in a series of recent reports, Dw2 was shown to be present in 50 to 70% of Caucasian MS patients and was far less frequent in controls (van den Berg-Loonen & Lucas 1975, Bertrams et al. 1975, Moller et al. 1975, Grosse-Wilde et al. 1976 Opelz et al. 1976). An extensive study of MS in Israel has been carried out by the same team of investigators since 1960 in order to analyze the influence of environmental and genetic factors on the risk of acquiring MS and on prognosis. Israel was chosen for this project because of its excellent medical facilities and detailed demographic data. Israel’s population is of heterogeneous origin and consists mainly of immigrants who originated from many different countries. Their original communities were often genetically isolated for centuries. It is not surprising, therefore, that Jewish groups have been found to differ from one another in various polymorphic gene frequencies and other traits (Cohen 1971). The diversity of ethnic groups with their highly varied cultures and life styles permits study of many factors which could affect risk of acquiring MS.One important result of the ongoing study of MS in Israel
was the demonstration that the prevalence rate of MS increased with the geographic latitude of the region of origin of the immigrants. Specifically, immigrants to Israel from Europe had much higher rates of MS than those from Afro-Asian countries. Native-born Israelis had a high prevalence rate of MS, like that observed in European immigrants, regardless of the country of origin of their parents (Leibowitz & Alter 1973). The aim of the present study was to determine histocompatibility determinants among MS patients in Israel and to reexamine the associations between HLA specificities and MS in ethnic groups which differ in MS prevalence. Materials and Methods Patients Patients with MS, in the present work, were identified in a nation-wide search which has been ongoing since 1960. The search has involved all hospitals, clinics and outpatient facilities in the country. In addition, inquiry among those physicians whose practices might include MS patients was carried out. Data were also collected from the National Neurological Disease Registry which has been maintained in Israel since 1969. This Registry is based on the discharge diagnoses of all hospitalized patients. By mandate, these discharge diagnoses must be submitted to the Central Bureau of Statistics. Patients tentatively identified as having “MS” or a clinically similar disorder were interviewed and examined by us and those patients who satisfied the standardized diagnostic criteria of Allison & Millar (1954) were retained. The present study included 197 patients who satisfied the most stringent of these clinical criteria. These patients gave a history of remissions
293
MS IN ISRAELI JEWISH PATIENTS
and exacerbations of neurological deficit attributable only to involvement of the central nervous system and, on examination, they had clinical evidence of scattered lesions of the neuraxis. The country of origin of the patients and their parents was determined at the time of the original interview. Forty-one patients were Jews of Mediterranean and Middle Eastern origin, 105, 11 and 28 were Jews originating from east, south and north-west Europe, respectively, and 12 were native-born Israelis. Controls Controls consisted of 455 individuals unrelated to the patients. They included unrelated blood donors, hospital personnel and volunteers whose parents, and in most cases parental grand-parents, had been born in the same country. One-hundred-andseventy-one were Mediterranean and Middle Eastern Jews, whereas 21 1, 19 and 38 were Jews originating from east, south and north-west Europe respectively, and 19 were native-born Israeli Jews.
Tissue Typing Lymphocytes were separated from defibrinated blood samples by Ficol-Hypaque flotation density equilibrium centrifugation (Boyum 1968). The fluororchromasia cytotoxicity test was employed for HLA typing of fresh and frozen (-180°C) lymphocytes (Bodmer et al. 1967). Each individual was typed at least twice for the following specificities: HLA-AI, 2, 3, 9, 10, 11, 28, 29, w19 and HLA-BS, 7, 8 , 12, 13, 14, 18, 27, w16, w17, w22, w35, w40. Generally, four to six doantisera were utilized to test for each specificity. Typing sera were obtained from the National Institutes of Health (NIH) serum bank, from other investigators and some
were obtained locally. Table 1 summarizes the sera utilized in this study for the identification of HLA-A3, B7 and Bw40. HLA-D Typing Lymphocytes for HLA-D typing were prepared from defibrinated blood and frozen in Jerusalem, Israel, using inactivated fetal calf serum with 5 % DMSO. The cell samples were kept overnight at -7OOC and transferred to liquid nitrogen until they were shipped in dry ice by air freight to &hus where typing was performed. The transportation time was normally 36 h and the viability of the lymphocytes was never below 90% as judged by trypan blue exclusion. The typing was carried out as previously described in detail (JBrgensen & Lamm 1974) using a battery of 10 different frozen homozygous typing cells (HTCs) of the following specificities: Dwl (2 HTCs), Dw2 (2 HTCs), Dw3 (3 HTCs), Dw4 (2 HTCs) and Dw6 (1 HTC). Some of these typing cells were defined during Table 1 H L A ANoantisera for the Identification of HLA-A3, B7 and Bw40 HLA Specificity A3 A3,All A3 A3
B7 B7 87, week A3 B7, Bw22 B7,B27 Bw40,B13 Bw40, B13 Bw21 Bw40, B7 Bw40,B7 Bw40 Bw40
Serum
Source
Kraska Storm Gallardo 854 P. Harris 37294168.9
NIH 2-65-0-10-21-01 NIH 2-50-6-03-22-02 NIH 2-50-7-01-17-01 Kissmeyer Payne Kissmeyer
Hasty Melnikoff Cowen Bras
Perkins Perkins Perkins Searle
Quinteros Wong Henke Walker McDaniel
Perkins Payne Perkins Perkins Grumet
294
BRAUTBAR ET AL.
the VI Histocompatibility Testing Workshop (Thorsby & Piazza 1975), while the remaining HTCs were compared with the workshop reference cells. In particular, one of the Dw2 cells was workshop cell 13-009. For the evaluation of the MLC typing experiments, normalization procedures were carried out both responderwise and stimulatorwise. First, relative responses were calculated as the percentage of the response in relation to the average response against three different pools, each of three cells. Second, stabilized relative responses (SRR) were calculated according to the 75 percentile criterion of Ryder et al. (1975). Finally the SRR was interpreted using the following rules: < 40% typing response meant antigen was present; 40-60% meant ambiguous response; > 60% positive MLC meant antigen not present. The antigen assignments depended on concurrent results with two or three typing cells except for Dw6, where only one HTC was available. Calculations Gene frequencies were estimated by a method devised by Nam & Gart (1976) which is based both on Bernstein’s method and on maximum likelihood procedures. The linkage disequilibrium parameter was calculated from the population data according to Mattiuz et al. (1970). The disease-HLA relationship has been expressed as the incidence ratio, i.e. the risk of contracting the disease for antigenpositive individuals relative to those without the antigen (Woolf 1955). In this communication, “relative risk” is used synonymously with “incidence ratio.” One special problem in the present study involves comparisons among pooled data from different ethnic groups which have
*,
different basic gene frequencies. This type of difficulty has been overcome by Woolf (1955) who elaborated a method for comparing risk figures from different populations and, if appropriate, for making a joint estimate of the incidence ratio from all studies. Woolf‘s paper also included a procedure for testing the significance of the deviation of the incidence ratio from 1. Table 2 GeographkYethnic grouping according to birthplace 1. Afro-Asia 2. South Europe (Yugoslavia, Bulgaria, Greece, Albania, Portugal, Spain, Italy) 3. East Europe (U.S.S.R.,Poland, Rumania, Czechoslovakia, Hungary). 4. West Europe (Germany, Austria, Switzerland) 5 . North Europe and America (Finland, Sweden Norway, Denmark, United Kingdom, Ireland, Holland, Belgium, Luxembourg, France, America, Canada). 6 . Native Israeli.
Results Because of limited numbers, it was necessary to group the subjects into those born in Afro-Asia, South Europe, East Europe, West or North Europe or America and Israel. Furthermore, to increase the genetic homogeneity of the Israeli-born subjects, they were grouped according to the birthplace of the father. I t has been shown that more than 80% of the fathers born outside Israel select a spouse from the same continent (Leibowitz & Alter 1973). The details of this geographic/ethnic grouping are given in Table 2. The basic HLA data are given in Tables 3 and 4. When the antigen frequencies in the five control samples were compared (x2 test with 4 d f ) it appeared that six of 23 antigens were heterogeneous at the 5% level while only three showed a significant heterogeneity at the 1% level. This was
1
MSC
CON
2 MS
P
-
0.54
11 16 41 171 5(46) 63(37) 19(46) l(6) 3 ~ 3 ) ~ 3 7 ) 5 ~ 1 ) 4(36) 2(18) 7(17) 6(38) 39(23) 56(33) 8(20) 5(31) 2(18) 23(14) 9(22) 0 4(36) 16(9) lO(24) 2(13) 0 57(33) 7(17) 9(56) 2(18) 4(io) 2(13) i(9) 13(8)
CONb
-
0.05 P ,. 0.79
28 8(29) 9(32) 2(7) 9(32) 11(39) 2(7) 8(29) 2(7)
38 9(24) 14(37) 9(24) 7(18) 9(24) 8(21) 8(21) 9(24)
105 15(14) 39(37) 18(17) 30(29) 32(31) 17(16) 32(31) 16(15)
211 63(30) 70(33) 30(14) 63(30) so(24) 22(10) 70(33) 21(10)
P
MS
4+5 CON
MS
3 CON
3(25) 305) 2(17) 7(58) 1(8)
8(42)
19
4(21) 2(11) 3(16) 4(21) 3(16)
MS
6(32) 7(37)
6 12 4(33) 1(8) 2(17)
CON
a) Grouped according to birthplace (as in Table 2) or, if native Israeli, according t o father's birthplace b) Control sample c) MSgroup P' P-value for joint incidence ratio being different from 1 P z P-value for different incidence ratios in the five ethnic groups P' P-value for heterogeneity of antigen frequencies in the five control groups P4 P-value for heterogeneity of antigen frequencies in control group 2, 3 , 4 f 5 and 6 3 P-value for Hardy-Weinberg distribution
HWS
No. = HLA-A1 HLA-A2 HLA-A3 HLA-A9 HLA-A10 HLA-A11 HLA-Awl9 HLA-A28
Ethnic Groupa
Table 3 Basic HLA-A and -B data no. (and %) of individuals with the antigen
n.s. -.02 -.04 n.s. n.s.
n.s. n.s. n.s.
P'
n.s. n.s. n.s. -.02 n.s.
-.007 n.s. n.s.
P2
-.04 n.s.
-.02
n.S. n.s.
n.s. n.s. -.03
P4
n.s. n.s. n.s.
-.02 n.s.
n.s. n.s.
-.05
P3
2
2
=! m
-u 9
E z
2
E
k
P
2
5:
P
-
0.07
CONb
1
For footnotessee Table 3
Hws
HLA-BIZ HLA-B13 HLA-B14 HLA-Bwl5 HLA-Bwl6 HLA-Bwl7 HLA-B18 HLA-Bw21 HLA-Bw22 HLA-B27 HLA-Bw35 HLA-Bw3 7 HLA-Bw40
nu-=
No. = HLA-B5 HLA-B7
Ethnic Groupa
MSC
CON
2 MS
P
-
3
-
MS
0.65
14(13)
0.42 P
14(7)
CON
3(8)
CON
MS
508)
4+5
16)
CON
6
Table 4 Basic HLA-A and -B data no. (and %) of individuals with the antigen
l(8)
MS
n.s. n.s.
-
n.s. n.s. -
-BOO8
n.s.
n.s.
n.s.
n.s. n.s.
n.s. ns.
n.s. n.s. n.s.
n.s.
n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. ns. n.s. n.s.
P'
n.s. n.s. n.s.
P'
n.s.
n.s.
n.s.
n.s. n.s. n.s. n.s.
n.s.
n.s. n.s. ns. n.s. n.s.
-.W5
n.s. n.s.
-.05
n.s. n.s. n.s.
P4
n.s. -.05 n.s.
n.s. -.003 n.s. -.OW03 -.02
-.05
-.01.
P3
F
m
? 3 ?!P
297
MS IN ISRAELI JEWISH PATIENTS
primarily due t o a difference between the Afro-Asian group and the others, 8 5 and B12 being higher and B14 lower in Jews of Afro-Asian origin. When the four groups from Europe, USA, and Israel were compared, only one antigen (Bw15) showed a conspicuous heterogeneity. The fit t o Hardy-Weinberg distribution seems reasonable in the groups considered large enough for this test. Only the Afro-Asian and the East-
European control groups were considered sufficiently large to give reliable data on haplotype frequencies and gametic associations. The results of these calculations are presented in Tables 5A and 5B. There was considerable difference between the two groups and only one haplotype (A10, Bw16) showed a significant positive A value in both groups. Taken as a whole, the HLA-antigen pattern in Jews was definitely Caucasian
Table SA Haplotype data for Jewish controls of different ethnic stock. Haplotypes with significant A in either of the populations A B 1, 8 1, 13 1, w17 1, w40 2, w14 2, 27 3.12 3, w14 9, w35 10,w14 10, w16 10, w17 11, 5 11, w35 11,w40 w19,13 28, w14 28, w22
Afro Asia ( N = 171) -202 2192.' 1326 1219 -0312. -307 2192. 12 -584 -573 1642.. -335 1372. 1842.. -69 -251 319 236
680 3209 2224 2024 -579 -170 3803 465 2406 -296 2132 -15 2057 2704 132 645 469 270
East Europe (N = 211) 1110. -435 4592*** -1233.. - 528 1080.. - 384 1707. 2657' -2157. 3392*** -2286.. 557 489 912.. 2084*** 1142. 535.
*level of significances: * = -0.05 * * = -0.01 First figure in each entry is A (X l o 5 ) Second F i r e is haplotype frequency (X 10')
1667 126 6194 -676 1629 1547 29 2599 5797 -637 4808 -1059 931 1522 1099 2717 1767 641
* * * = -0.001
Table S B Haplotype data for Jewish controls of different ethnic stock hhplotypes with highest frequency or delta value Afro Asia (N = 171) ~-
A3, B12
Highest Frequency
A2, B5 A l , Bw35
Largest delta
A3, B12 A l , B13 A l l , Bw35
East Europe (N = 211) ~~
A l , Bw17 A9, Bw35 A10, Bw16 A l , Bw17
A10,Bw16 A9, Bw35
298
BRAUTBAR ET AL.
Table 6 Incidence ratios for M S and Bw40 Bw40
Population
MS
Afro Asia 21.9%of 41 South Europe 9.0%of 1 1 East Europe 13.3%of 105 N+WEur.+ USA 17.8%of 28 Native Israeli 8.3%of 12 Combined risk Heterogeneity among risks
95%LIM
7.6%of 171 6.2%of 16 6.6%of 211
1.34 0.08 0.09
< 3.14 < 8.67 < 1 . 5 0 < 26.85 < 2.16 < 4.72
7.8%of 38 5.2%of 19
0.55 0.09
< 2.53 < 11.65 1.43077 < 1.63 < 28.90 0.11299 < 2.51 < 4.30 11.30117
1.46
Risk
95%LIM
xa
CONT
PROB.
6.69628 0.00966 0.07587 0.78296 3.75350 0.05269 0.23164 0.73676
0.00077 0.76825 0.94265
(Bodmer et al. 1972), and the data from borderline significantly increased relative Israel reported here agreed closely with risks. Neither A3 nor B7 gave any evidence findings in other East-Asiatic Caucasians at all of association with MS. A1 may have (Turks, Lebanese and Arabs) and also been connected with different risks in various populations: increased in Afroin other Asiatic Caucasians. With respect to the haplotype pat- Asians and in South, West and North tern, neither of the typical North-West Europeans, and decreased in East EuroEuropean haplotypes ( A l , B8 nor A3, peans and native Israelis. Awl9 may also B7) showed positive linkage disequilibrium have indicated different risks in the various in the Jews whom we studied. This result populations. Table 7 shows the outcome of HLA-D is consistent with a similar lack of increase of these haplotypes in other East- typing of 28 patients and 22 controls. ern European Caucasians. Furthermore, Only one of the controls and one of the the only “typical” Jewish haplotype patients could be assigned a Dw2 speci(A10, Bw16) which was increased in ficity and, as expected, both were B7 East-European and Afro-Asian Jews, has positive. Six individuals were Dw3 posialso been identified as increased in other tive (two patients, four controls) and four Caucasian groups including Turks, Basques, of these were also B8 positive. islanders from the Hebrides in Scotland, Discussion and in France. The genetic heterogeneity of the Jewish When Woolf‘s (1955) test was applied people who have lived in small isolated t o the data for these five ethnic groups groups for centuries affects interpretation (Tables 3 and 4) only one antigen (Bw40) of the results. If the gene frequencies are showed a highly significant ( P < 0.001) very different in various Jewish ethnic positive association with MS. Furthermore, stocks, then even small biases in the selecthe incidence ratio for this antigen seemed t o be the same in all five groups, and Table 7 HLA-D antigen assignments in Jewish M S based on the joint data the risk of a Bw40patients and controls positive individual t o contract MS was 2.5 Dwl 2 3 4 6 times the risk of a Bw4O-negative indiMS (N=28) 1 1 2 2 3 vidual (Table 6). 0 1 4 0 0 A10 and A l l were both associated with Controls (N= 22)
M S IN ISRAELI JEWISH PATIENTS
299
tion of controls may give rise to spurious been HLA-D typed, but the finding that associations between a disease and HLA only one of 28 Jewish patients could be antigens. The present study, however, has assigned the Dw2 specificity precludes demonstrated typical Middle-East Caucas- any association of Dw2 with MS in these ian HLA-A and -B patterns in all ethnic/ patients of a similar magnitude to that geographic groups studied, and only a observed in Caucasian and American few antigens (B5, B12, B14, and Bw15) Negro patients. In Danish, Norwegian, gave significant evidence at the 1% level and South-German MS patients, Dw2 in the various groups. In particular, Bw40 was found in 60, 72 and 44%, respectshowed none and A3 and B7 only border- ively, compared to approximately 20% line significance (P < 0.02) heterogeneity. of controls, which gives an average risk In this context it must be remembered of 5.3 (Jersild et al. 1975). In 31 Amerthat when 23 antigens are considered, ican-Negro patients, Dw2 was found in then the true 5% and 1%significance levels 35%, while it could not be detected among correspond to P-values of 0.05/23 = 34 controls (Dupont et al. 1977). 0.002 and 0.01/23 = 0.0004, respectively. The lack of association in Jewish MS These considerations weaken the evidence patients with any Dw specificities tested of heterogeneity. for ( w l , 2, 3, 4, 6) does not necessarily I t must be concluded from the present mean that the HLA-D genes are not assocstudy that neither A3 nor B7 was assoc- iated with MS among Jews. Another D iated with MS in Jews. Instead, Bw40 specificity which was not detected by the showed an increase from 5 to 8% in the available HTCs may be implicated. different control groups compared with 8 The difference between the findings in to 22% in the various MS groups. The rela- Jews, Negroes and Caucasians could, tive risk in individuals with Bw40 seemed perhaps, best be explained by assuming to be the same in all ethnic/geographical that the same MS disease susceptibility groups and was estimated at 2.5. The (MSS) gene is operating in all three populastatistical significance of this risk corre- tions, but that it is found in associations sponds to a true 0.02 level. with different alleles at the HLA region The published studies on MS and HLA in each. In that case, Dw2 per se could in peopIe of Caucasian stock have uni- not be the MSS gene. Alternatively, it formly shown highly significant associa- could also be postulated that different tions with A3 and B7 with joint incidence MSS genes are operating in the three ratios of 1.3 and 1.6, respectively (Jersild populations, viz. Dw2 in European Caucet al. 1975). In contrast, it has been shown asians and Negroes and some as yet unithat MS in American Negroes was assoc- dentified gene in Jews. Different disease iated with Bw40 with a relative risk of susceptibility genes could arise if differabout 3 while MS was not significantly ent pathogenic agents were responsible associated with A3 or B7 (Dupont et al. for MS in different geographic areas. 1977). The association of Bw40 with MS It will therefore be interesting to see in Jews was perhaps surprising as Jews whether Jewish MS patients living in have an HLA profile much more similar Europe or America have Dw2 or not. It must, however, be observed that to European Caucasians than to Negroes. a remarkable difference exists in the Only a few Jewish individuals have
3 00
BRAUTBAR ET AL.
haplotype profile of the Afro-Asian and East-European groups (Tables 5A and 5B). Only one haplotype (A10, Bw16) showed a similar positive linkage disequilibrium in both groups, and the relevant antigen, Bw40, seems to be involved in two significant associations, positively with A1 1 and negatively with Al, but this could only be detected in the East-European group and not among the Afro-Asians. This might mean that the same MSS gene cannot be associated with MS in all Jewish subgroups unless the MSS is very closely linked to or identical with Bw40. A close linkage to an HLA-B gene would not be compatible with the finding in European Caucasians and American Negroes that the MSS gene seems to map closer to the HLA-D than to the HLA-B locus (Jersild et al. 1975). But it must be stressed that a closer association of the MSS gene with HLA-D than with HLA-B does not necessarily mean that this gene maps closer to HLA-D. The haplotype data must, however, be accepted with reservations, for if sampling has occurred from heterogenous subpopulations, then spurious associations may well arise. One important rationale for the study of MS in the Israeli groups was the finding t h a t ' the prevalence rate of MS among Israeli-born descendants of Afro-Asian immigrants was comparable to the high prevalence rate among Israeli-born descendants of immigrants from the European continent, and not as low as the prevalence of MS rate in the Afro-Asian immigrants per se. As Israel resembles European countries in life-style more than it does underdeveloped Afro-Asian countries, the high rate of MS in Afro-Asians born in Israel was interpreted to mean that environmental factors, e.g. the socio-economic status o r life-style, were more impor-
tant than genetic factors in determining a high MS prevalence rate. The present study demonstrates the existence of a genetic factor for MS which is common to the various ethnic groups. This factor, however, cannot explain the observed differences in prevalence rates as Bw40 has similar frequencies in ethnic groups with high and low rates of MS. But it must be admitted that the sample sizes are too small definitely to rule out the existence of such differences in Bw40 frequencies. Only one antigen (B14) showed a significant decrease - from 34% in the north-western to 8% in the south-eastern populations - paralleling the decreasing MS prevalence rate from north to south across Europe and Afro-Asia. However, the B14 antigen did not seem to be associated with MS in Jews. Therefore, an environmental rather than a genetic factor (e.g. HLA) still seems to be a more likely explanation for differences in MS frequency in different latitudes. References Acheson, E. D. (1972) The Epidemiology of Multiple Sclerosis. Multiple Sclerosis: A Reappraisal. pp. 42-80. Baltimore, The Williams and Wilkins Company. Allison, R. S. & Millar, J . H. D. (1954) R e valence and familial incidence of disseminated sclerosis: report to the Northern Ireland Hospitals Authority on the results of a threeyear survey. Ulster med. J . 23 (suppl. 2), 1-92. Amason, B. G . W., Fuller, T. C., Lehrich, J. R. & Wray, S. H. (1974) Histocompatibility types and measles antibodies in multiple sclerosis and optic neuritis. J . neurol. Sci. 22,419-428. Berg-Loonen E . M. van den & Lucas, K. J . (1975) LD7a typing in 4 6 patients with multiple sclerosis. Histocompatibility Testing 1975, pp. 773-777. Munksgaard, Copenhagen.
MS IN ISRAELI JEWISH PATIENTS Berry, R. J. (1969) Genetical factors in the etiology of multiple sclerosis. Acta. Neurol. 45,459-483. Bertrams, J. & Kuwert, E. (1972) HL-A antigen frequencies in multiple sclerosis. Significant increase of HL-A3, HL-A10 and W5, and decrease of HL-A12. European Neurology 7, 74-78. Bertrams, J., Grosse-Wilde, H., Netzel, B., Mempel, W. & Kuwert, E. (1975) SD-HL-A3-7LD Pi(7a): A Caucasian histocompatibility super haplotype with increased susceptibility to multiple sclerosis. Histocompatibility Testing 1975, pp. 782-787, Munksgaard, Copenhagen. Bodmer, J., Rocques, P., Bodmer, W., Colombani, J., Dagos, L., Dausset, J. & Piazza, A. (1972) Joint Report of the Fifth International Histocompatibility Workshop. Histocompatibility Testing 1972a, pp. 621-667, Ed.: Dausset, J. & Colombani, J. (Munksgaard, Copenhagen). Bodmer, W., Tripp, M. & Bodmer, J. (1967) Application of the fluorochromatic cytotoxicity assay to human leucocyte typing. Histocompatibility Testing 1967, pp. 341350. Munksgaard, Copenhagen. Boyum, A. (1968) Separation of leukocytes from blood and bone marrow. Scand. J. Clin. Lab. Invest. Suppl. 97. Cohen, T. (1971) Genetic markers in migrants t o Israel. Israel J. med. Sci. 7, 1509-1514. Dupont, B., Jersild, C., Hausen, G. A., Nielsen, L. S., Thomson, M. & Svejgaard, A. (1973) Typing for MLC determinants by means of LD-homozygous and LD heterozygous test cells. Transplant. Proc. 5, 1543-1549. Dupont, B., Lisak, R. P., Jenild, C., Hansen, J. A., Siberberg, D. H., Whitsett, C., Zwieman, B. & Ciogoli, K. (1977) HLA antigens in black American patients with multiple sclerosis. Transpl. Proc. In Press. Gross-Wilde, H., Bertrams, J., Netzel, B. & Kuwert, E. K. (1976) Frequency of four HLA-D alleles in multiple sclerosis patients. Abstract in: First Internaitonal Symposium on HLA and disease, p. 67. Jersild, C., DUpont, B., Fog, T., Platz, P. J. & Svejgaard, A. (1975) Histocompatibility determinants in multiple sclerosis. Transp. Rev. 22, 148-163.
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Jersild, C., Fog, T., Hausen, G. S., Thomsen, M., Svjgaard, A. & Dupont, B. (1973b) Histocompatibility determinants in multiple sclerosis with special reference to the clinical course. Lancet 2, 1221-1224. Jersild, C., Svjgaard, A., Fog, T. & Ammitzbdl, T. (1973a) HL-A antigens and disease. I. Multiple Sclerosis. Tissue Antigens 3, 243250. Jdrgensen, F. & Lamm, L. U. (1974) MLC- a micro-modification of the mixed leucocyte culture technique. Tissue Antigens 4, 482492,1974. Jdrgensen, F., Lamm, L. V. & Kissmeyer-Nielsen, F. (1973) Mixed lymphocyte culture with inbred individuals. An approach t o MLC typing. Tissue Antigens 3, 323-339. Leibowitz, U. & Alter, M. (1973) Multiple Sclerosis: clues to its causes. Amsterdam, North Holland Publishing Company. Mattiuz, P. L., Ihdo, D., Piazza, A., Ceppelini, R. & Bodmer, W. F. (1970) New Approaches to the Population Genetic and Segregation Analysis of the HL-A System. Histocompatibility Testing, pp. 193-205. Munksgaard, Copenhagen. Moller, E., Link, H., Matell, G., Olhzgen, B. & Stendahl, L. (1975) LD alleles in ankylosing spondylities, myasthenia gravis, multiple sclerosis and optic neuritis. Histocompatibility Testing 1975, pp. 778-781, Munksgaard, Copenhagen. Naito, S., Namerow, N., Mickey, M. R. & Terasaki, P. I. (1972) Multiple Sclerosis: Association with HL-A3. Tissue Antigens 2, 1-4. Nam, J. -M. & Cart, J. J. (1976) Bernsteins and gene-counting methods in generalized ABOlike systems. Ann. Hum. Genet. London. 39, 361. Opelz, G., Terasaki, P., Myers, L., Ellison, G., Ebers, J., Zabriski, H., Kempe, H., Sibley, W. & Sever, J. (1976) HLA-D locus typing in 350 patients with multiple sclerosis. Abstract in: First International Symposium on HLA and Disease, p. 76. Ryder, L. P., Thomsen, M.,Platz, P. & Svejgaard, A. (1975) Data reduction in LD-typing. Histocompatibility Testing 1975, p. 557, ed. Kissmeyer-Nielsen, F. Munksgaard, Copenhagen. Thorsby, E. & Piazza, A. (1975) Joint report
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from the Sixth International Histocompatibility Workshop Conference 11. typing for HLA-D (LD-1or MLC) determinants. Histocompatibility Testing, 1975. ed. KissmeyerNielsen, F. Munksgaard, Copenhagen. Woolf, B. (1955) On estimating the relation between blood group and disease. Ann. Hum. Genet. 19,251-253.
Address: Chaim Brautbar Department of Immunology Hadassah Hebrew University Medical Center Jerusalem Israel
Histocompatibility Determinants in Israeli Jewish Patients with Multiple Sclerosis Chaim Brautbar' , isac Cohen' , Esther Kahana', Milton Alter3, Fritz Jbrgensen4 and Lars Lamm4 Department of Immunology and Laboratory of Immunohematology, Hadassah Hebrew University Medical Center, Jerusalem, Israel; Department of Neurology, Hadassah Hebrew University Medical Center, Jerusalem, Israel; Department of Neurology, Temple University Health Sciences Center, Philadelphia, U.S.A. and Blood Grouping and Tissue Typing Laboratory, The University Hospital, Arhus, Denmark
'
The distribution of 24 HLA antigens of the A and B loci was investigated in 197 Israeli Jewish patients with multiple sclerosis (MS) from various Jewish ethnic origins including central and eastern Europe, countries bordering the Mediterranean, the Middle East and from native-born Israelis. The results were compared with the HLA antigen frequencies in a control sample of 455 unrelated individuals representing the general Jewish population. The frequency of HLA-Bw40 among all MS patients (15%) was significantly greater (P < 0.001) than among the controls (7%). In contrast to the findings in MS patients from other populations, there was no increased frequency of A3 and 07 and Dw2 was present in only one out of 28 patients. The study showed a similar distribution of HLA-A and -B locus antigens, especially of Bw40, in Jews of diverse ethnic origins represented in the control group.
Received for publication 1 1 March, revised, accepted 26 April 1977
Although an environmental factor has been suggested as the most likely cause of multiple sclerosis (MS) (Acheson 1972, Leibowitz & Alter 1973) the disease occurs 12 to 20 times more frequently within the family of a patient than in the general population (Berry 1969). Therefore, a genetic factor may also play a role in etiology. Numerous studies have shown as association between MS and certain histocom-
patibility (HLA) determinants, notably HLA-A3 of the first segregant series and B7 of the second series, (Bertram & Kuwert 1972, Naito et al. 1972, Jersaid et al. 1973a, Arnason et al. 1974). In a recent study it was clearly shown that HLA-B7 is the antigen with the greatest overall frequency (Jersild et al. 1975). Since it became possible to define and characterize lymphocytedefined determinants utilizing HLA-D homozygous typing
This work was supported, in part, by the Kroc Foundation for Medical Research, the Danish Medical Research Council, the Israel Multiple Sclerosis Society and Minktry of Health, and b y a special research grant kindly donated by Mr. and Mrs. Forrest Shaklee.
29 2
BRAUTBAR ET AL.
cells (Dupont et al. 1973, Jbrgensen et al. 1973) an HLA-B7 associated D locus determinant has been sought which was even more closely associated with MS. Indeed, Jersild et al. (1973b) were able to show a higher frequency of the lymphocyte-defined determinant Dw2 (formerly LD-7a) in MS patients (70%) than in the general population (16%). The presence of Dw2 was also apparently associated with a more malignant course of the disease (Jersild et al. 1973b). Further studies have confirmed that the frequency of Dw2 among MS patients is significantly higher than HLA-B7 (Jersild et al. 1975). In fact, in a series of recent reports, Dw2 was shown to be present in 50 to 70% of Caucasian MS patients and was far less frequent in controls (van den Berg-Loonen & Lucas 1975, Bertrams et al. 1975, Moller et al. 1975, Grosse-Wilde et al. 1976 Opelz et al. 1976). An extensive study of MS in Israel has been carried out by the same team of investigators since 1960 in order to analyze the influence of environmental and genetic factors on the risk of acquiring MS and on prognosis. Israel was chosen for this project because of its excellent medical facilities and detailed demographic data. Israel’s population is of heterogeneous origin and consists mainly of immigrants who originated from many different countries. Their original communities were often genetically isolated for centuries. It is not surprising, therefore, that Jewish groups have been found to differ from one another in various polymorphic gene frequencies and other traits (Cohen 1971). The diversity of ethnic groups with their highly varied cultures and life styles permits study of many factors which could affect risk of acquiring MS.One important result of the ongoing study of MS in Israel
was the demonstration that the prevalence rate of MS increased with the geographic latitude of the region of origin of the immigrants. Specifically, immigrants to Israel from Europe had much higher rates of MS than those from Afro-Asian countries. Native-born Israelis had a high prevalence rate of MS, like that observed in European immigrants, regardless of the country of origin of their parents (Leibowitz & Alter 1973). The aim of the present study was to determine histocompatibility determinants among MS patients in Israel and to reexamine the associations between HLA specificities and MS in ethnic groups which differ in MS prevalence. Materials and Methods Patients Patients with MS, in the present work, were identified in a nation-wide search which has been ongoing since 1960. The search has involved all hospitals, clinics and outpatient facilities in the country. In addition, inquiry among those physicians whose practices might include MS patients was carried out. Data were also collected from the National Neurological Disease Registry which has been maintained in Israel since 1969. This Registry is based on the discharge diagnoses of all hospitalized patients. By mandate, these discharge diagnoses must be submitted to the Central Bureau of Statistics. Patients tentatively identified as having “MS” or a clinically similar disorder were interviewed and examined by us and those patients who satisfied the standardized diagnostic criteria of Allison & Millar (1954) were retained. The present study included 197 patients who satisfied the most stringent of these clinical criteria. These patients gave a history of remissions
293
MS IN ISRAELI JEWISH PATIENTS
and exacerbations of neurological deficit attributable only to involvement of the central nervous system and, on examination, they had clinical evidence of scattered lesions of the neuraxis. The country of origin of the patients and their parents was determined at the time of the original interview. Forty-one patients were Jews of Mediterranean and Middle Eastern origin, 105, 11 and 28 were Jews originating from east, south and north-west Europe, respectively, and 12 were native-born Israelis. Controls Controls consisted of 455 individuals unrelated to the patients. They included unrelated blood donors, hospital personnel and volunteers whose parents, and in most cases parental grand-parents, had been born in the same country. One-hundred-andseventy-one were Mediterranean and Middle Eastern Jews, whereas 21 1, 19 and 38 were Jews originating from east, south and north-west Europe respectively, and 19 were native-born Israeli Jews.
Tissue Typing Lymphocytes were separated from defibrinated blood samples by Ficol-Hypaque flotation density equilibrium centrifugation (Boyum 1968). The fluororchromasia cytotoxicity test was employed for HLA typing of fresh and frozen (-180°C) lymphocytes (Bodmer et al. 1967). Each individual was typed at least twice for the following specificities: HLA-AI, 2, 3, 9, 10, 11, 28, 29, w19 and HLA-BS, 7, 8 , 12, 13, 14, 18, 27, w16, w17, w22, w35, w40. Generally, four to six doantisera were utilized to test for each specificity. Typing sera were obtained from the National Institutes of Health (NIH) serum bank, from other investigators and some
were obtained locally. Table 1 summarizes the sera utilized in this study for the identification of HLA-A3, B7 and Bw40. HLA-D Typing Lymphocytes for HLA-D typing were prepared from defibrinated blood and frozen in Jerusalem, Israel, using inactivated fetal calf serum with 5 % DMSO. The cell samples were kept overnight at -7OOC and transferred to liquid nitrogen until they were shipped in dry ice by air freight to &hus where typing was performed. The transportation time was normally 36 h and the viability of the lymphocytes was never below 90% as judged by trypan blue exclusion. The typing was carried out as previously described in detail (JBrgensen & Lamm 1974) using a battery of 10 different frozen homozygous typing cells (HTCs) of the following specificities: Dwl (2 HTCs), Dw2 (2 HTCs), Dw3 (3 HTCs), Dw4 (2 HTCs) and Dw6 (1 HTC). Some of these typing cells were defined during Table 1 H L A ANoantisera for the Identification of HLA-A3, B7 and Bw40 HLA Specificity A3 A3,All A3 A3
B7 B7 87, week A3 B7, Bw22 B7,B27 Bw40,B13 Bw40, B13 Bw21 Bw40, B7 Bw40,B7 Bw40 Bw40
Serum
Source
Kraska Storm Gallardo 854 P. Harris 37294168.9
NIH 2-65-0-10-21-01 NIH 2-50-6-03-22-02 NIH 2-50-7-01-17-01 Kissmeyer Payne Kissmeyer
Hasty Melnikoff Cowen Bras
Perkins Perkins Perkins Searle
Quinteros Wong Henke Walker McDaniel
Perkins Payne Perkins Perkins Grumet
294
BRAUTBAR ET AL.
the VI Histocompatibility Testing Workshop (Thorsby & Piazza 1975), while the remaining HTCs were compared with the workshop reference cells. In particular, one of the Dw2 cells was workshop cell 13-009. For the evaluation of the MLC typing experiments, normalization procedures were carried out both responderwise and stimulatorwise. First, relative responses were calculated as the percentage of the response in relation to the average response against three different pools, each of three cells. Second, stabilized relative responses (SRR) were calculated according to the 75 percentile criterion of Ryder et al. (1975). Finally the SRR was interpreted using the following rules: < 40% typing response meant antigen was present; 40-60% meant ambiguous response; > 60% positive MLC meant antigen not present. The antigen assignments depended on concurrent results with two or three typing cells except for Dw6, where only one HTC was available. Calculations Gene frequencies were estimated by a method devised by Nam & Gart (1976) which is based both on Bernstein’s method and on maximum likelihood procedures. The linkage disequilibrium parameter was calculated from the population data according to Mattiuz et al. (1970). The disease-HLA relationship has been expressed as the incidence ratio, i.e. the risk of contracting the disease for antigenpositive individuals relative to those without the antigen (Woolf 1955). In this communication, “relative risk” is used synonymously with “incidence ratio.” One special problem in the present study involves comparisons among pooled data from different ethnic groups which have
*,
different basic gene frequencies. This type of difficulty has been overcome by Woolf (1955) who elaborated a method for comparing risk figures from different populations and, if appropriate, for making a joint estimate of the incidence ratio from all studies. Woolf‘s paper also included a procedure for testing the significance of the deviation of the incidence ratio from 1. Table 2 GeographkYethnic grouping according to birthplace 1. Afro-Asia 2. South Europe (Yugoslavia, Bulgaria, Greece, Albania, Portugal, Spain, Italy) 3. East Europe (U.S.S.R.,Poland, Rumania, Czechoslovakia, Hungary). 4. West Europe (Germany, Austria, Switzerland) 5 . North Europe and America (Finland, Sweden Norway, Denmark, United Kingdom, Ireland, Holland, Belgium, Luxembourg, France, America, Canada). 6 . Native Israeli.
Results Because of limited numbers, it was necessary to group the subjects into those born in Afro-Asia, South Europe, East Europe, West or North Europe or America and Israel. Furthermore, to increase the genetic homogeneity of the Israeli-born subjects, they were grouped according to the birthplace of the father. I t has been shown that more than 80% of the fathers born outside Israel select a spouse from the same continent (Leibowitz & Alter 1973). The details of this geographic/ethnic grouping are given in Table 2. The basic HLA data are given in Tables 3 and 4. When the antigen frequencies in the five control samples were compared (x2 test with 4 d f ) it appeared that six of 23 antigens were heterogeneous at the 5% level while only three showed a significant heterogeneity at the 1% level. This was
1
MSC
CON
2 MS
P
-
0.54
11 16 41 171 5(46) 63(37) 19(46) l(6) 3 ~ 3 ) ~ 3 7 ) 5 ~ 1 ) 4(36) 2(18) 7(17) 6(38) 39(23) 56(33) 8(20) 5(31) 2(18) 23(14) 9(22) 0 4(36) 16(9) lO(24) 2(13) 0 57(33) 7(17) 9(56) 2(18) 4(io) 2(13) i(9) 13(8)
CONb
-
0.05 P ,. 0.79
28 8(29) 9(32) 2(7) 9(32) 11(39) 2(7) 8(29) 2(7)
38 9(24) 14(37) 9(24) 7(18) 9(24) 8(21) 8(21) 9(24)
105 15(14) 39(37) 18(17) 30(29) 32(31) 17(16) 32(31) 16(15)
211 63(30) 70(33) 30(14) 63(30) so(24) 22(10) 70(33) 21(10)
P
MS
4+5 CON
MS
3 CON
3(25) 305) 2(17) 7(58) 1(8)
8(42)
19
4(21) 2(11) 3(16) 4(21) 3(16)
MS
6(32) 7(37)
6 12 4(33) 1(8) 2(17)
CON
a) Grouped according to birthplace (as in Table 2) or, if native Israeli, according t o father's birthplace b) Control sample c) MSgroup P' P-value for joint incidence ratio being different from 1 P z P-value for different incidence ratios in the five ethnic groups P' P-value for heterogeneity of antigen frequencies in the five control groups P4 P-value for heterogeneity of antigen frequencies in control group 2, 3 , 4 f 5 and 6 3 P-value for Hardy-Weinberg distribution
HWS
No. = HLA-A1 HLA-A2 HLA-A3 HLA-A9 HLA-A10 HLA-A11 HLA-Awl9 HLA-A28
Ethnic Groupa
Table 3 Basic HLA-A and -B data no. (and %) of individuals with the antigen
n.s. -.02 -.04 n.s. n.s.
n.s. n.s. n.s.
P'
n.s. n.s. n.s. -.02 n.s.
-.007 n.s. n.s.
P2
-.04 n.s.
-.02
n.S. n.s.
n.s. n.s. -.03
P4
n.s. n.s. n.s.
-.02 n.s.
n.s. n.s.
-.05
P3
2
2
=! m
-u 9
E z
2
E
k
P
2
5:
P
-
0.07
CONb
1
For footnotessee Table 3
Hws
HLA-BIZ HLA-B13 HLA-B14 HLA-Bwl5 HLA-Bwl6 HLA-Bwl7 HLA-B18 HLA-Bw21 HLA-Bw22 HLA-B27 HLA-Bw35 HLA-Bw3 7 HLA-Bw40
nu-=
No. = HLA-B5 HLA-B7
Ethnic Groupa
MSC
CON
2 MS
P
-
3
-
MS
0.65
14(13)
0.42 P
14(7)
CON
3(8)
CON
MS
508)
4+5
16)
CON
6
Table 4 Basic HLA-A and -B data no. (and %) of individuals with the antigen
l(8)
MS
n.s. n.s.
-
n.s. n.s. -
-BOO8
n.s.
n.s.
n.s.
n.s. n.s.
n.s. ns.
n.s. n.s. n.s.
n.s.
n.s. n.s. n.s. n.s. n.s. n.s. n.s. n.s. ns. n.s. n.s.
P'
n.s. n.s. n.s.
P'
n.s.
n.s.
n.s.
n.s. n.s. n.s. n.s.
n.s.
n.s. n.s. ns. n.s. n.s.
-.W5
n.s. n.s.
-.05
n.s. n.s. n.s.
P4
n.s. -.05 n.s.
n.s. -.003 n.s. -.OW03 -.02
-.05
-.01.
P3
F
m
? 3 ?!P
297
MS IN ISRAELI JEWISH PATIENTS
primarily due t o a difference between the Afro-Asian group and the others, 8 5 and B12 being higher and B14 lower in Jews of Afro-Asian origin. When the four groups from Europe, USA, and Israel were compared, only one antigen (Bw15) showed a conspicuous heterogeneity. The fit t o Hardy-Weinberg distribution seems reasonable in the groups considered large enough for this test. Only the Afro-Asian and the East-
European control groups were considered sufficiently large to give reliable data on haplotype frequencies and gametic associations. The results of these calculations are presented in Tables 5A and 5B. There was considerable difference between the two groups and only one haplotype (A10, Bw16) showed a significant positive A value in both groups. Taken as a whole, the HLA-antigen pattern in Jews was definitely Caucasian
Table SA Haplotype data for Jewish controls of different ethnic stock. Haplotypes with significant A in either of the populations A B 1, 8 1, 13 1, w17 1, w40 2, w14 2, 27 3.12 3, w14 9, w35 10,w14 10, w16 10, w17 11, 5 11, w35 11,w40 w19,13 28, w14 28, w22
Afro Asia ( N = 171) -202 2192.' 1326 1219 -0312. -307 2192. 12 -584 -573 1642.. -335 1372. 1842.. -69 -251 319 236
680 3209 2224 2024 -579 -170 3803 465 2406 -296 2132 -15 2057 2704 132 645 469 270
East Europe (N = 211) 1110. -435 4592*** -1233.. - 528 1080.. - 384 1707. 2657' -2157. 3392*** -2286.. 557 489 912.. 2084*** 1142. 535.
*level of significances: * = -0.05 * * = -0.01 First figure in each entry is A (X l o 5 ) Second F i r e is haplotype frequency (X 10')
1667 126 6194 -676 1629 1547 29 2599 5797 -637 4808 -1059 931 1522 1099 2717 1767 641
* * * = -0.001
Table S B Haplotype data for Jewish controls of different ethnic stock hhplotypes with highest frequency or delta value Afro Asia (N = 171) ~-
A3, B12
Highest Frequency
A2, B5 A l , Bw35
Largest delta
A3, B12 A l , B13 A l l , Bw35
East Europe (N = 211) ~~
A l , Bw17 A9, Bw35 A10, Bw16 A l , Bw17
A10,Bw16 A9, Bw35
298
BRAUTBAR ET AL.
Table 6 Incidence ratios for M S and Bw40 Bw40
Population
MS
Afro Asia 21.9%of 41 South Europe 9.0%of 1 1 East Europe 13.3%of 105 N+WEur.+ USA 17.8%of 28 Native Israeli 8.3%of 12 Combined risk Heterogeneity among risks
95%LIM
7.6%of 171 6.2%of 16 6.6%of 211
1.34 0.08 0.09
< 3.14 < 8.67 < 1 . 5 0 < 26.85 < 2.16 < 4.72
7.8%of 38 5.2%of 19
0.55 0.09
< 2.53 < 11.65 1.43077 < 1.63 < 28.90 0.11299 < 2.51 < 4.30 11.30117
1.46
Risk
95%LIM
xa
CONT
PROB.
6.69628 0.00966 0.07587 0.78296 3.75350 0.05269 0.23164 0.73676
0.00077 0.76825 0.94265
(Bodmer et al. 1972), and the data from borderline significantly increased relative Israel reported here agreed closely with risks. Neither A3 nor B7 gave any evidence findings in other East-Asiatic Caucasians at all of association with MS. A1 may have (Turks, Lebanese and Arabs) and also been connected with different risks in various populations: increased in Afroin other Asiatic Caucasians. With respect to the haplotype pat- Asians and in South, West and North tern, neither of the typical North-West Europeans, and decreased in East EuroEuropean haplotypes ( A l , B8 nor A3, peans and native Israelis. Awl9 may also B7) showed positive linkage disequilibrium have indicated different risks in the various in the Jews whom we studied. This result populations. Table 7 shows the outcome of HLA-D is consistent with a similar lack of increase of these haplotypes in other East- typing of 28 patients and 22 controls. ern European Caucasians. Furthermore, Only one of the controls and one of the the only “typical” Jewish haplotype patients could be assigned a Dw2 speci(A10, Bw16) which was increased in ficity and, as expected, both were B7 East-European and Afro-Asian Jews, has positive. Six individuals were Dw3 posialso been identified as increased in other tive (two patients, four controls) and four Caucasian groups including Turks, Basques, of these were also B8 positive. islanders from the Hebrides in Scotland, Discussion and in France. The genetic heterogeneity of the Jewish When Woolf‘s (1955) test was applied people who have lived in small isolated t o the data for these five ethnic groups groups for centuries affects interpretation (Tables 3 and 4) only one antigen (Bw40) of the results. If the gene frequencies are showed a highly significant ( P < 0.001) very different in various Jewish ethnic positive association with MS. Furthermore, stocks, then even small biases in the selecthe incidence ratio for this antigen seemed t o be the same in all five groups, and Table 7 HLA-D antigen assignments in Jewish M S based on the joint data the risk of a Bw40patients and controls positive individual t o contract MS was 2.5 Dwl 2 3 4 6 times the risk of a Bw4O-negative indiMS (N=28) 1 1 2 2 3 vidual (Table 6). 0 1 4 0 0 A10 and A l l were both associated with Controls (N= 22)
M S IN ISRAELI JEWISH PATIENTS
299
tion of controls may give rise to spurious been HLA-D typed, but the finding that associations between a disease and HLA only one of 28 Jewish patients could be antigens. The present study, however, has assigned the Dw2 specificity precludes demonstrated typical Middle-East Caucas- any association of Dw2 with MS in these ian HLA-A and -B patterns in all ethnic/ patients of a similar magnitude to that geographic groups studied, and only a observed in Caucasian and American few antigens (B5, B12, B14, and Bw15) Negro patients. In Danish, Norwegian, gave significant evidence at the 1% level and South-German MS patients, Dw2 in the various groups. In particular, Bw40 was found in 60, 72 and 44%, respectshowed none and A3 and B7 only border- ively, compared to approximately 20% line significance (P < 0.02) heterogeneity. of controls, which gives an average risk In this context it must be remembered of 5.3 (Jersild et al. 1975). In 31 Amerthat when 23 antigens are considered, ican-Negro patients, Dw2 was found in then the true 5% and 1%significance levels 35%, while it could not be detected among correspond to P-values of 0.05/23 = 34 controls (Dupont et al. 1977). 0.002 and 0.01/23 = 0.0004, respectively. The lack of association in Jewish MS These considerations weaken the evidence patients with any Dw specificities tested of heterogeneity. for ( w l , 2, 3, 4, 6) does not necessarily I t must be concluded from the present mean that the HLA-D genes are not assocstudy that neither A3 nor B7 was assoc- iated with MS among Jews. Another D iated with MS in Jews. Instead, Bw40 specificity which was not detected by the showed an increase from 5 to 8% in the available HTCs may be implicated. different control groups compared with 8 The difference between the findings in to 22% in the various MS groups. The rela- Jews, Negroes and Caucasians could, tive risk in individuals with Bw40 seemed perhaps, best be explained by assuming to be the same in all ethnic/geographical that the same MS disease susceptibility groups and was estimated at 2.5. The (MSS) gene is operating in all three populastatistical significance of this risk corre- tions, but that it is found in associations sponds to a true 0.02 level. with different alleles at the HLA region The published studies on MS and HLA in each. In that case, Dw2 per se could in peopIe of Caucasian stock have uni- not be the MSS gene. Alternatively, it formly shown highly significant associa- could also be postulated that different tions with A3 and B7 with joint incidence MSS genes are operating in the three ratios of 1.3 and 1.6, respectively (Jersild populations, viz. Dw2 in European Caucet al. 1975). In contrast, it has been shown asians and Negroes and some as yet unithat MS in American Negroes was assoc- dentified gene in Jews. Different disease iated with Bw40 with a relative risk of susceptibility genes could arise if differabout 3 while MS was not significantly ent pathogenic agents were responsible associated with A3 or B7 (Dupont et al. for MS in different geographic areas. 1977). The association of Bw40 with MS It will therefore be interesting to see in Jews was perhaps surprising as Jews whether Jewish MS patients living in have an HLA profile much more similar Europe or America have Dw2 or not. It must, however, be observed that to European Caucasians than to Negroes. a remarkable difference exists in the Only a few Jewish individuals have
3 00
BRAUTBAR ET AL.
haplotype profile of the Afro-Asian and East-European groups (Tables 5A and 5B). Only one haplotype (A10, Bw16) showed a similar positive linkage disequilibrium in both groups, and the relevant antigen, Bw40, seems to be involved in two significant associations, positively with A1 1 and negatively with Al, but this could only be detected in the East-European group and not among the Afro-Asians. This might mean that the same MSS gene cannot be associated with MS in all Jewish subgroups unless the MSS is very closely linked to or identical with Bw40. A close linkage to an HLA-B gene would not be compatible with the finding in European Caucasians and American Negroes that the MSS gene seems to map closer to the HLA-D than to the HLA-B locus (Jersild et al. 1975). But it must be stressed that a closer association of the MSS gene with HLA-D than with HLA-B does not necessarily mean that this gene maps closer to HLA-D. The haplotype data must, however, be accepted with reservations, for if sampling has occurred from heterogenous subpopulations, then spurious associations may well arise. One important rationale for the study of MS in the Israeli groups was the finding t h a t ' the prevalence rate of MS among Israeli-born descendants of Afro-Asian immigrants was comparable to the high prevalence rate among Israeli-born descendants of immigrants from the European continent, and not as low as the prevalence of MS rate in the Afro-Asian immigrants per se. As Israel resembles European countries in life-style more than it does underdeveloped Afro-Asian countries, the high rate of MS in Afro-Asians born in Israel was interpreted to mean that environmental factors, e.g. the socio-economic status o r life-style, were more impor-
tant than genetic factors in determining a high MS prevalence rate. The present study demonstrates the existence of a genetic factor for MS which is common to the various ethnic groups. This factor, however, cannot explain the observed differences in prevalence rates as Bw40 has similar frequencies in ethnic groups with high and low rates of MS. But it must be admitted that the sample sizes are too small definitely to rule out the existence of such differences in Bw40 frequencies. Only one antigen (B14) showed a significant decrease - from 34% in the north-western to 8% in the south-eastern populations - paralleling the decreasing MS prevalence rate from north to south across Europe and Afro-Asia. However, the B14 antigen did not seem to be associated with MS in Jews. Therefore, an environmental rather than a genetic factor (e.g. HLA) still seems to be a more likely explanation for differences in MS frequency in different latitudes. References Acheson, E. D. (1972) The Epidemiology of Multiple Sclerosis. Multiple Sclerosis: A Reappraisal. pp. 42-80. Baltimore, The Williams and Wilkins Company. Allison, R. S. & Millar, J . H. D. (1954) R e valence and familial incidence of disseminated sclerosis: report to the Northern Ireland Hospitals Authority on the results of a threeyear survey. Ulster med. J . 23 (suppl. 2), 1-92. Amason, B. G . W., Fuller, T. C., Lehrich, J. R. & Wray, S. H. (1974) Histocompatibility types and measles antibodies in multiple sclerosis and optic neuritis. J . neurol. Sci. 22,419-428. Berg-Loonen E . M. van den & Lucas, K. J . (1975) LD7a typing in 4 6 patients with multiple sclerosis. Histocompatibility Testing 1975, pp. 773-777. Munksgaard, Copenhagen.
MS IN ISRAELI JEWISH PATIENTS Berry, R. J. (1969) Genetical factors in the etiology of multiple sclerosis. Acta. Neurol. 45,459-483. Bertrams, J. & Kuwert, E. (1972) HL-A antigen frequencies in multiple sclerosis. Significant increase of HL-A3, HL-A10 and W5, and decrease of HL-A12. European Neurology 7, 74-78. Bertrams, J., Grosse-Wilde, H., Netzel, B., Mempel, W. & Kuwert, E. (1975) SD-HL-A3-7LD Pi(7a): A Caucasian histocompatibility super haplotype with increased susceptibility to multiple sclerosis. Histocompatibility Testing 1975, pp. 782-787, Munksgaard, Copenhagen. Bodmer, J., Rocques, P., Bodmer, W., Colombani, J., Dagos, L., Dausset, J. & Piazza, A. (1972) Joint Report of the Fifth International Histocompatibility Workshop. Histocompatibility Testing 1972a, pp. 621-667, Ed.: Dausset, J. & Colombani, J. (Munksgaard, Copenhagen). Bodmer, W., Tripp, M. & Bodmer, J. (1967) Application of the fluorochromatic cytotoxicity assay to human leucocyte typing. Histocompatibility Testing 1967, pp. 341350. Munksgaard, Copenhagen. Boyum, A. (1968) Separation of leukocytes from blood and bone marrow. Scand. J. Clin. Lab. Invest. Suppl. 97. Cohen, T. (1971) Genetic markers in migrants t o Israel. Israel J. med. Sci. 7, 1509-1514. Dupont, B., Jersild, C., Hausen, G. A., Nielsen, L. S., Thomson, M. & Svejgaard, A. (1973) Typing for MLC determinants by means of LD-homozygous and LD heterozygous test cells. Transplant. Proc. 5, 1543-1549. Dupont, B., Lisak, R. P., Jenild, C., Hansen, J. A., Siberberg, D. H., Whitsett, C., Zwieman, B. & Ciogoli, K. (1977) HLA antigens in black American patients with multiple sclerosis. Transpl. Proc. In Press. Gross-Wilde, H., Bertrams, J., Netzel, B. & Kuwert, E. K. (1976) Frequency of four HLA-D alleles in multiple sclerosis patients. Abstract in: First Internaitonal Symposium on HLA and disease, p. 67. Jersild, C., DUpont, B., Fog, T., Platz, P. J. & Svejgaard, A. (1975) Histocompatibility determinants in multiple sclerosis. Transp. Rev. 22, 148-163.
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Jersild, C., Fog, T., Hausen, G. S., Thomsen, M., Svjgaard, A. & Dupont, B. (1973b) Histocompatibility determinants in multiple sclerosis with special reference to the clinical course. Lancet 2, 1221-1224. Jersild, C., Svjgaard, A., Fog, T. & Ammitzbdl, T. (1973a) HL-A antigens and disease. I. Multiple Sclerosis. Tissue Antigens 3, 243250. Jdrgensen, F. & Lamm, L. U. (1974) MLC- a micro-modification of the mixed leucocyte culture technique. Tissue Antigens 4, 482492,1974. Jdrgensen, F., Lamm, L. V. & Kissmeyer-Nielsen, F. (1973) Mixed lymphocyte culture with inbred individuals. An approach t o MLC typing. Tissue Antigens 3, 323-339. Leibowitz, U. & Alter, M. (1973) Multiple Sclerosis: clues to its causes. Amsterdam, North Holland Publishing Company. Mattiuz, P. L., Ihdo, D., Piazza, A., Ceppelini, R. & Bodmer, W. F. (1970) New Approaches to the Population Genetic and Segregation Analysis of the HL-A System. Histocompatibility Testing, pp. 193-205. Munksgaard, Copenhagen. Moller, E., Link, H., Matell, G., Olhzgen, B. & Stendahl, L. (1975) LD alleles in ankylosing spondylities, myasthenia gravis, multiple sclerosis and optic neuritis. Histocompatibility Testing 1975, pp. 778-781, Munksgaard, Copenhagen. Naito, S., Namerow, N., Mickey, M. R. & Terasaki, P. I. (1972) Multiple Sclerosis: Association with HL-A3. Tissue Antigens 2, 1-4. Nam, J. -M. & Cart, J. J. (1976) Bernsteins and gene-counting methods in generalized ABOlike systems. Ann. Hum. Genet. London. 39, 361. Opelz, G., Terasaki, P., Myers, L., Ellison, G., Ebers, J., Zabriski, H., Kempe, H., Sibley, W. & Sever, J. (1976) HLA-D locus typing in 350 patients with multiple sclerosis. Abstract in: First International Symposium on HLA and Disease, p. 76. Ryder, L. P., Thomsen, M.,Platz, P. & Svejgaard, A. (1975) Data reduction in LD-typing. Histocompatibility Testing 1975, p. 557, ed. Kissmeyer-Nielsen, F. Munksgaard, Copenhagen. Thorsby, E. & Piazza, A. (1975) Joint report
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from the Sixth International Histocompatibility Workshop Conference 11. typing for HLA-D (LD-1or MLC) determinants. Histocompatibility Testing, 1975. ed. KissmeyerNielsen, F. Munksgaard, Copenhagen. Woolf, B. (1955) On estimating the relation between blood group and disease. Ann. Hum. Genet. 19,251-253.
Address: Chaim Brautbar Department of Immunology Hadassah Hebrew University Medical Center Jerusalem Israel
